Protein oxidative modifications: new mass spectrometry approaches to detection in biological samples

Corinne M. Spickett, E. Silajdzic, L. Mouls, K. Thomson, Andrew Pitt

Research output: Contribution to conferenceAbstract

Abstract

In inflammatory diseases, release of oxidants leads to oxidative damage to proteins. The precise nature of oxidative damage to individual proteins depends on the oxidant involved. Chlorination and nitration are markers of modification by the myeloperoxidase-H2O2-Cl- system and nitric oxide-derived oxidants, respectively. Although these modifications can be detected by western blotting, currently no reliable method exists to identify the specific sites damage to individual proteins in complex mixtures such as clinical samples. We are developing novel LCMS2 and precursor ion scanning methods to address this. LC-MS2 allows separation of peptides and detection of mass changes in oxidized residues on fragmentation of the peptides. We have identified indicative fragment ions for chlorotyrosine, nitrotyrosine, hydroxytyrosine and hydroxytryptophan. A nano-LC/MS3 method involving the dissociation of immonium ions to give specific fragments for the oxidized
residues has been developed to overcome the problem of false positives from ions isobaric to these immonium ions that exist in unmodified peptides. The approach has proved able to identify precise protein modifications in individual proteins and mixtures of proteins. An alternative methodology involves multiple reaction monitoring for precursors and fragment ions are specific to oxidized and chlorinated proteins, and this has been tested with human serum albumin. Our ultimate aim is to apply this methodology to the detection of oxidative post-translational modifications in clinical samples for disease diagnosis, monitoring the outcomes of therapy, and improved understanding of disease biochemistry.
LanguageEnglish
Pages43
Number of pages1
Publication statusPublished - 2009
EventSFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention - Roma, Italy
Duration: 27 Sep 2009 → …

Conference

ConferenceSFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention
CountryItaly
CityRoma
Period27/09/09 → …

Fingerprint

Mass spectrometry
Ions
Oxidants
Proteins
Peptides
Nitration
5-Hydroxytryptophan
Biochemistry
Monitoring
Chlorination
Complex Mixtures
Serum Albumin
Peroxidase
Nitric Oxide
Scanning

Bibliographical note

Abstract published online in Abstracts, Free Radical Research, 43(s1) p.S43.

Keywords

  • mass spectrometry
  • oxidants
  • oxidative damage
  • protein
  • protein modifications
  • Science (General)

Cite this

Spickett, C. M., Silajdzic, E., Mouls, L., Thomson, K., & Pitt, A. (2009). Protein oxidative modifications: new mass spectrometry approaches to detection in biological samples. 43. Abstract from SFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention, Roma, Italy.
Spickett, Corinne M. ; Silajdzic, E. ; Mouls, L. ; Thomson, K. ; Pitt, Andrew. / Protein oxidative modifications : new mass spectrometry approaches to detection in biological samples. Abstract from SFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention, Roma, Italy.1 p.
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Spickett, CM, Silajdzic, E, Mouls, L, Thomson, K & Pitt, A 2009, 'Protein oxidative modifications: new mass spectrometry approaches to detection in biological samples' SFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention, Roma, Italy, 27/09/09, pp. 43.

Protein oxidative modifications : new mass spectrometry approaches to detection in biological samples. / Spickett, Corinne M.; Silajdzic, E.; Mouls, L.; Thomson, K.; Pitt, Andrew.

2009. 43 Abstract from SFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention, Roma, Italy.

Research output: Contribution to conferenceAbstract

TY - CONF

T1 - Protein oxidative modifications

T2 - new mass spectrometry approaches to detection in biological samples

AU - Spickett, Corinne M.

AU - Silajdzic, E.

AU - Mouls, L.

AU - Thomson, K.

AU - Pitt, Andrew

N1 - Abstract published online in Abstracts, Free Radical Research, 43(s1) p.S43.

PY - 2009

Y1 - 2009

N2 - In inflammatory diseases, release of oxidants leads to oxidative damage to proteins. The precise nature of oxidative damage to individual proteins depends on the oxidant involved. Chlorination and nitration are markers of modification by the myeloperoxidase-H2O2-Cl- system and nitric oxide-derived oxidants, respectively. Although these modifications can be detected by western blotting, currently no reliable method exists to identify the specific sites damage to individual proteins in complex mixtures such as clinical samples. We are developing novel LCMS2 and precursor ion scanning methods to address this. LC-MS2 allows separation of peptides and detection of mass changes in oxidized residues on fragmentation of the peptides. We have identified indicative fragment ions for chlorotyrosine, nitrotyrosine, hydroxytyrosine and hydroxytryptophan. A nano-LC/MS3 method involving the dissociation of immonium ions to give specific fragments for the oxidizedresidues has been developed to overcome the problem of false positives from ions isobaric to these immonium ions that exist in unmodified peptides. The approach has proved able to identify precise protein modifications in individual proteins and mixtures of proteins. An alternative methodology involves multiple reaction monitoring for precursors and fragment ions are specific to oxidized and chlorinated proteins, and this has been tested with human serum albumin. Our ultimate aim is to apply this methodology to the detection of oxidative post-translational modifications in clinical samples for disease diagnosis, monitoring the outcomes of therapy, and improved understanding of disease biochemistry.

AB - In inflammatory diseases, release of oxidants leads to oxidative damage to proteins. The precise nature of oxidative damage to individual proteins depends on the oxidant involved. Chlorination and nitration are markers of modification by the myeloperoxidase-H2O2-Cl- system and nitric oxide-derived oxidants, respectively. Although these modifications can be detected by western blotting, currently no reliable method exists to identify the specific sites damage to individual proteins in complex mixtures such as clinical samples. We are developing novel LCMS2 and precursor ion scanning methods to address this. LC-MS2 allows separation of peptides and detection of mass changes in oxidized residues on fragmentation of the peptides. We have identified indicative fragment ions for chlorotyrosine, nitrotyrosine, hydroxytyrosine and hydroxytryptophan. A nano-LC/MS3 method involving the dissociation of immonium ions to give specific fragments for the oxidizedresidues has been developed to overcome the problem of false positives from ions isobaric to these immonium ions that exist in unmodified peptides. The approach has proved able to identify precise protein modifications in individual proteins and mixtures of proteins. An alternative methodology involves multiple reaction monitoring for precursors and fragment ions are specific to oxidized and chlorinated proteins, and this has been tested with human serum albumin. Our ultimate aim is to apply this methodology to the detection of oxidative post-translational modifications in clinical samples for disease diagnosis, monitoring the outcomes of therapy, and improved understanding of disease biochemistry.

KW - mass spectrometry

KW - oxidants

KW - oxidative damage

KW - protein

KW - protein modifications

KW - Science (General)

M3 - Abstract

SP - 43

ER -

Spickett CM, Silajdzic E, Mouls L, Thomson K, Pitt A. Protein oxidative modifications: new mass spectrometry approaches to detection in biological samples. 2009. Abstract from SFRR Europe Meeting 2009 Free radicals, Health and Lifestyle: from cell signalling to disease prevention, Roma, Italy.